Target recognition, RNA methylation activity and transcriptional regulation of the Dictyostelium discoideum Dnmt2-homologue (DnmA)

Please always quote using this URN: urn:nbn:de:bvb:20-opus-123149
  • Although the DNA methyltransferase 2 family is highly conserved during evolution and recent reports suggested a dual specificity with stronger activity on transfer RNA (tRNA) than DNA substrates, the biological function is still obscure. We show that the Dictyostelium discoideum Dnmt2-homologue DnmA is an active tRNA methyltransferase that modifies C38 in \(tRNA^{Asp(GUC)}\) in vitro and in vivo. By an ultraviolet-crosslinking and immunoprecipitation approach, we identified further DnmA targets. This revealed specific tRNA fragments bound byAlthough the DNA methyltransferase 2 family is highly conserved during evolution and recent reports suggested a dual specificity with stronger activity on transfer RNA (tRNA) than DNA substrates, the biological function is still obscure. We show that the Dictyostelium discoideum Dnmt2-homologue DnmA is an active tRNA methyltransferase that modifies C38 in \(tRNA^{Asp(GUC)}\) in vitro and in vivo. By an ultraviolet-crosslinking and immunoprecipitation approach, we identified further DnmA targets. This revealed specific tRNA fragments bound by the enzyme and identified \(tRNA^{Glu(CUC/UUC)}\) and \(tRNA^{Gly(GCC)}\) as new but weaker substrates for both human Dnmt2 and DnmA in vitro but apparently not in vivo. Dnmt2 enzymes form transient covalent complexes with their substrates. The dynamics of complex formation and complex resolution reflect methylation efficiency in vitro. Quantitative PCR analyses revealed alterations in dnmA expression during development, cell cycle and in response to temperature stress. However, dnmA expression only partially correlated with tRNA methylation in vivo. Strikingly, dnmA expression in the laboratory strain AX2 was significantly lower than in the NC4 parent strain. As expression levels and binding of DnmA to a target in vivo are apparently not necessarily accompanied by methylation, we propose an additional biological function of DnmA apart from methylation.show moreshow less

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Metadaten
Author: Sara Müller, Indra M. Windhof, Vladimir Maximov, Tomasz Jurkowski, Albert Jeltsch, Konrad U. Förstner, Cynthia M. Sharma, Ralph Gräf, Wolfgang Nellen
URN:urn:nbn:de:bvb:20-opus-123149
Document Type:Journal article
Faculties:Medizinische Fakultät / Institut für Molekulare Infektionsbiologie
Language:English
Parent Title (English):Nucleic Acids Research
ISSN:1362-4962
Year of Completion:2013
Volume:41
Issue:18
Pagenumber:8615-8627
Source:Nucleic Acids Research (2013) 41 (18): 8615-8627. doi: 10.1093/nar/gkt634
DOI:https://doi.org/10.1093/nar/gkt634
Dewey Decimal Classification:5 Naturwissenschaften und Mathematik / 57 Biowissenschaften; Biologie / 572 Biochemie
Tag:DNA methylferase homolog; TRNA(ASP); binding; drospophila; mechanism
Release Date:2016/03/02
Licence (German):License LogoCC BY-NC: Creative-Commons-Lizenz: Namensnennung, Nicht kommerziell